The present invention relates to intracellular receptors, methods for the modulation thereof, and methods for the identification of novel ligands therefor. In a particular aspect, the present invention relates to methods for the identification of compounds which function as ligands (or ligand precursors) for intracellular receptors. In another aspect, the present invention relates to novel chimeric constructs and uses therefor.
A central problem in eukaryotic molecular biology continues to be the elucidation of molecules and mechanisms that mediate specific gene regulation. As part of the scientific attack on this problem, a great deal of work has been done in efforts to identify ligands (i.e., exogenous inducers) which are capable of mediating specific gene regulation. Additional work has been done in efforts to identify other molecules involved in specific gene regulation.
Although much remains to be learned about the specifics of gene regulation, it is known that ligands modulate gene transcription by acting in concert with intracellular components, including intracellular receptors and discrete DNA sequences known as hormone response elements (HREs).
The identification of compounds which directly or indirectly interact with intracellular receptors, and thereby affect transcription of hormone-responsive genes, would be of significant value, e.g., for therapeutic applications.
Transcriptional silencing mediated by nuclear receptors plays an important role in development, cell differentiation, and is directly linked to the oncogenic activity of v-erbA. The mechanism underlying this effect is unknown but is one key to understanding the molecular basis of hormone action. Accordingly, the identification of components involved in transcriptional silencing would represent a great advance in current understanding of mechanisms that mediate specific gene regulation.
Other information helpful in the understanding and practice of the present invention can be found In commonly assigned U.S. Pat. Nos. 5,071,773 and 4,981,784; and U.S. patent application Ser. No. 326,240, filed Mar. 17, 1389 (now issued as U.S. Pat. No. 5,597,693, based on continuation in part application Ser. No. 494,618, filed Mar. 16, 1990); Ser. No. 370,407, filed Jun. 22, 1989 (now issued as U.S. Pat. No. 5,260,432) and Ser. No. 438,757, filed Nov. 16, 1989 (now issued as U.S. Pat. No. 5,091,518), all of which are hereby incorporated herein by reference in their entirety.
In accordance with the present invention, we have discovered a novel receptor interacting factor, referred to herein as xe2x80x9cSMRTxe2x80x9d, i.e., a silencing mediator (co-repressor) for retinoic acid receptor (RAR) and thyroid hormone receptor (TR). SMRT is a novel protein whose association with RAR and TR both in solution and on DNA response elements is destabilized by ligand. The interaction of SMRT with mutant receptors correlates with the transcriptional silencing activities of receptors.
In vivo, SMRT functions as a potent co-repressor. A GAL4 DNA binding domain (DBD) fusion of SMRT behaves as a frank repressor of a GAL4-dependent reporter. Together, these data identify a novel class of cofactor which is believed to represent an important mediator of hormone action.